Method for demethylating target methylated region of genome DNA

Abstract

It is an object of the present invention to provide a demethylation method that, from among multiple methylated regions present in the genome DNA of a living cell, can demethylate a specified methylated region only without influencing the other methylated regions, and to achieve the object, a target methylated region present in either a sense or antisense DNA chain in genome DNA of a living cell is demethylated, by artificially generating in the living cell the following (a) RNA comprising a base sequence equivalent to the base sequence of a region comprising all or part of the target methylated region in the DNA chain where the target methylated region is present, or (b) RNA comprising a base sequence equivalent to the base sequence of a region comprising a region neighboring the target methylated region in the DNA chain where the target methylated region is present.

1 . A method for demethylating a target methylated region present in either sense or antisense DNA chain in genome DNA of a living cell, the method comprising a step of artificially generating in the living cell the following RNA (a) or (b), 
 (a) RNA comprising a base sequence equivalent to the base sequence of a region comprising all or part of the target methylated region in the DNA chain where the target methylated region is present;    (b) RNA comprising a base sequence equivalent to the base sequence of a region comprising a region neighboring the target methylated region in the DNA chain where the target methylated region is present.    
     
     
         2 . The method according to  claim 1 , further comprising a step of introducing into the living cell a recombinant vector that can express the RNA (a) or (b).  
     
     
         3 . A recombinant vector that can express the RNA (a) or (b) according to  claim 1 .  
     
     
         4 . A demethylation kit for demethylating a target methylated region present in either sense or antisense DNA chain in genome DNA of a living cell, the demethylation kit comprising the recombinant vector according to  claim 3 .  
     
     
         5 . A living cell into which the recombinant vector according to  claim 3  has been introduced.  
     
     
         6 . A tissue or organ produced by culturing the living cell according to  claim 5 .  
     
     
         7 . A cell nucleus isolated from the living cell according to  claim 5 .  
     
     
         8 . A cloned animal produced by using the cell nucleus according to  claim 7 .  
     
     
         9 . A method for producing a tissue or organ by culturing a live cell, comprising a step of demethylating a target methylated region present in either sense or antisense DNA chain in genome DNA of a living cell by artificially generating in the living cell the RNA (a) or (b) according to  claim 1 .  
     
     
         10 . A method for producing a cloned animal using the cell nucleus according to  claim 7  as the cell nucleus to implant in an enucleated unfertilized egg.